Genetic Engineering

Lesson Objectives At the end of this lesson you should be able to Define Genetic Engineering Understand that GE alters DNA Understand the function of restriction enzymes Be able to explain isolation, cutting , insertion , transformation and expression Discuss three applications of GE Discuss the ethical issues of genetic engineering

Genetic Engineering The simple addition, deletion, or manipulation of a single trait in an organism to create a desired change.

Genetic Engineering What you need to know Manipulation and alteration of genes Three applications: one plant, one animal, one micro-organism Process involving isolation, transformation, and expression

Genetic Engineering Is: Artificially copying a piece of DNA from one organism and joining this copy of DNA into the DNA of another organism Modern biotechnology relies on newer techniques, such as genetic engineering, to incorporate genetic material from one living organism into another. The DNA is cut from the donor cell and inserted into the DNA of another organism (vector) e.g. bacterium. The bacterium will then manufacture the protein coded for by the gene. Products of biotechnology include medications, human insulin, and enzymes used in laundry detergents and cheese-making. More recently, the use of biotechnology has led to new pesticide products that control a variety of pests. www.clipartguide.com

Purpose of Genetic Engineering It allows genes from one organism to be inserted into a cell of a different organism of a different species. Examples: Human genes can be inserted into a bacterium Human genes can be inserted into cells from other animals Bacterium genes can be inserted into plant cells

Genetic Engineering Genetic engineering means that DNA from different organisms can be combined Bacteria can be engineered to produce human proteins Human genes can be inserted into other animals

Genetic Engineering - Overview The altered DNA is called recombinant DNA Recombinant DNA is joined to other unrelated DNA in the organism This is called gene splicing. - tiny segments of a gene are taken out and replaced by different genes

Learning Check What is Genetic Engineering? What is the purpose of Genetic Reengineering? Give three examples of Genetic Engineering What is the difference between altered and recombinant DNA? What is meant by gene splicing?

Transgenic Organisms Organisms altered by genetic engineering. Genetic material changed by other than random natural breeding Gene transfer -moving a gene from one organism to another.

What Transgenic means 'Trans-' means 'crossing from one place to another‘ The '-genic' bit means genes So it means that bits of genes from different living things have been bolted together and spliced into another organism to make a new one which does something which the scientists want it to do.

Examples of Transgenic Organisms GMO- genetically modified organism GEO-genetically enhanced organism For example Plants that resists a particular type of weed killer Sheep which makes some special substance in its milk.

Learning Check What the word transgenic mean? What is a transgenic organism? Give examples of transgenic organisms

5 Stages involved in GE Isolation Cutting Ligation and Insertion Transformation Expression

1. Isolation (a) Isolation of a specific gene from donor e.g. human Cells broken open Genetic probe added Reveals position of the gene of interest The first step involves breaking open the cells of the donor to release the DNA and isolate the gene of interest e.g. insulin producing gene. Cells are broken open using chemicals and enzymes e.g. washing up liquid Donor DNA is extracted Genetic probe is added A DNA probe consists of a small fragment of DNA labelled with an enzyme, a radioactive tag or a flurescent dye tag. The probe will bind to a complementary DNA sequence by base pairing. Identifying the presence and location of the gene of interest Genetic probe Position of gene of interest Donor DNA

1. Isolation (b) Isolation of plasmid from a bacterial cell The DNA from the bactterial cell is released. A bacterial cell contains a circular loop of DNA called a plasmid. The plasmid is isolated from the bacterial cell. The plasmid will act as a vector for carrying a new gene i.e. the gene from the donor will be inserted into the plasmid DNA.. www.sci.sdsu.edu

2. Cutting Restriction enzymes act as molecular scissors and cut DNA at specific sites called restriction sites Restriction site Restriction ezymes The donor DNA and plasmid DNA are cut using enzymes called restriction enzymes. Restriction enzymes recognise specific sequence of bases Act as a molecular scissors to cut the DNA strand within the recognition sequence. The donor DNA and plasmid DNA are cut using the same restriction enzymes. Bacteria use restriction enzymes to defend themselves against attacks from bacterial viruses. The enzymes cut invading viral DNA and render it harmless. Clipartguide.com

2. Cutting Restriction site Donor DNA Plasmid Restriction enzymes © Biology Support Service 2007

Cutting Donor DNA Sticky Ends Plasmid Restriction enzymes cut the DNA from the donor at specific points The cut ends have sticky ends The same restriction enzymes cut the DNA from the plasmid also revealing sticky ends As a result of cutting by the restriction enzymes the donor DNA and the plasmid are left with unpaired bases known as sticky ends.

DNA Ligase http://www.slic2.wsu.edu:82/hurlbert/micro101/pages/Chap10.html#Sticky_ended_cut Ligation –rejoining cut fragments of DNA and forming artificial recombinant molecules

Ligation and Insertion

Recombinant DNA introduced into bacterial cell 4. Transformation Recombinant DNA introduced into bacterial cell Bacterial chromosome Bacterial cell Recombinant DNA

5. Expression Bacterial cell reproduces by Binary Fisson Bacterial cell produces the polypeptide Coded for by the donor DNA Universality of genetic code Plasmid will produce the polypeptide coded for by the donor DNA

Summary of steps Donor DNA 1. Cut with restriction enzymes Donor DNA Plasmid 1. Cut with restriction enzymes Donor DNA Sticky Ends 2. Ligase bonds sticky ends together Recombinant DNA

Expression Expression is getting the organism with the recombinant DNA to produce the desired protein When the protein is produced in large amounts it is isolated and purified

Learning Check What are the stages involved in GE? Outline what happens in the stage isolation ? What enzyme is used to cut the DNA? Can you explain what happens in transformation? What is meant by gene expression?

Animals used in GE The human gene to clot blood has been inserted into the DNA of sheep Sheep produce human clotting factor needed for Haemophiliacs in their milk Goats produce a protein to treat emphysema

Applications (Micro-organisms) Production of humulin Insulin is a hormone that controls sugar levels in an organism. Diabetes occurs in people when there is too little or too much insulin produced. To control diabetes, sufferers usually inject insulin once or twice daily. Until the mid-1980's most insulin was produced by extracting a human-equivalent insulin from the pancreas of animals (usually pigs). GM insulin ( Humilin) is a genetically modified form of insulin. The genetic sequence for insulin production is removed from human DNA. This is then inserted into the DNA of a bacteria, Escherrichia coli (E. coli). The gene inserted into the bacteria cell is inherited from cell to cell as the cells multiply. The insulin protein is produced (expressed) by the cells. The insulin is then extracted from the cells.  Problems with BSE in Britain have made users wary of products derived from animals. GM insulin carries none of these concerns. GM insulin is also cheaper to produce than pig-based extraction. During the production process there are points at which contamination of the insulin mix can occur. This caused initial problems with patient reactions to these contaminants. Some patients have reported hypoglycaemic problems after switching to GM insulin - they have lost the ability to detect the onset of hypoglycaemia. However, various studies indicate that over a third of long-term diabetics lose this ability after 10 years of treatment.   Used by diabetics

Plant Application Golden Rice – a possible solution to Vitamin A deficiency. I INSERTED ‘POSSIBLE’ BEFORE ‘SOLUTION’ HERE.

Applications of Genetic Engineering

Gene Therapy It involves modifying human DNA either to repair it or to replace a faulty gene. The idea of gene therapy is to overcome the effects of a mutation which causes a genetic disease. Cystic fibrosis is the best known disease where gene therapy has been tried.

Diagnostic Tests Genetic engineering can produce very specific and sensitive diagnostic tests for many diseases, using engineered proteins. This new technology is also opening up novel ways of delivering medicines to specific targets.

Vaccines Genetically engineered microbes can be used to produce the antigens needed in a safe and controllable way. The use of genetically modified yeast cells to produce a vaccine against the hepatitis B virus has been a major success story.

Plants Weedkiller resistant crops - Weeds die but the crops survive Vitamin A in Rice - The gene which produces vitamin A was taken from daffodils and put into rice to help prevent blindness

Micro organisms Bacteria can make human insulin This prevented many diabetics from getting an allergic reaction to animal insulin Bacteria make interferon which can fight virus infections and some cancers

Learning Check Can you outline 3 uses of Genetic Engineering? What is the purpose of Pharming? Outline a use for Gene Therapy What have GE modified yeast cells been used to produce? What do you feel are the ethical issues associated with GE?

What have you learned? Can you………… Define Genetic Engineering Understand that GE alters DNA Understand the function of restriction enzymes Be able to explain isolation, cutting , insertion , transformation and expression Discuss three applications of GE Discuss the ethical issues of genetic engineering

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